Two-stage dynamic deregulation of metabolism improves process robustness & scalability in engineered E. coli

We report that two-stage dynamic control improves bioprocess robustness as a result of the dynamic deregulation of central metabolism. Dynamic control is implemented during stationary phase using combinations of CRISPR interference and controlled proteolysis to reduce levels of central metabolic enzymes. Reducing the levels of key enzymes alters metabolite pools resulting in deregulation of the metabolic network. Deregulated networks are less sensitive to environmental conditions improving process robustness. Process robustness in turn leads to predictable scalability, minimizing the need for traditional process optimization. We validate process robustness and scalability of strains and bioprocesses synthesizing the important industrial chemicals alanine, citramalate and xylitol. Predictive high throughput approaches that translate to larger scales are critical for metabolic engineering programs to truly take advantage of the rapidly increasing throughput and decreasing costs of synthetic biology. •Two-stage dynamic metabolic control enables deregulation of central metabolism.•Deregulation of the metabolic network improves process robustness.•Process robustness improves scalability from 96 well plates to instrumented bioreactors.•Rapid scale up from lab bioreactors to pilot bioreactors.